tsA mutants of SV40 greatly overproduce early mRNA when growth at low temperature is followed by a temperature shift. This early mRNA will be mapped by electron microscopy and sized accurately, to determine how it is transcribed and processed. Comparable experiments with late SV40 mRNA are also planned, under conditions where processing steps may have been altered. Mutant hamster cell lines that overproduce the first three enzymes of de novo pyrimidine nucleotide synthesis by about 100-fold will be used as a source of mRNA for these enzymes, which appear to be parts of a single polypeptide. Following its isolation and characterization by translation in vitro, the mRNA will be used to synthesize labeled cDNA, which will be used as a probe of possible mechanisms responsible for the observed change in gene regulation. BIBLIOGRAPHIC REFERENCES: R. K. Johnson, T. Inouye, A. Goldin and G. R. Stark. Antitumor Activity of N-Phosphonacetyl-L-Aspartate, a Transition State Analog Inhibitor of Aspartate Transcarbamylase, Cancer Res. 36, 2720 (1976). G. R. Stark, Multifunctional proteins: one gene-more than one enzyme. Trends in Biochemical Sciences (1977), in press.